The orphan nuclear receptor steroidogenic factor 1 (SF-1) plays essential roles in endocrine development and function. SF-1 knockout (KO) mice exhibit adrenal and gonadal agenesis, impaired pituitary gonadotrope function, and marked structural abnormalities of the ventromedial hypothalamic nucleus (VMH). The proposed studies seek to expand our understanding of these pleiotropic roles of SF-1, focusing specifically on its roles in gonadal development. First, we will use the Cre-loxP technology to inactivate SF-1 in specific cell lineages of the testes and ovaries. By using different promoters to direct Cre expression, we envision that we will achieve both cell-type specific inactivation as well as inactivation at different stages of development. We also will use a transgenic enhanced green fluorescent protein (eGFP) reporter gene directed by SF-1 regulatory sequences to purify SF-l-expressing cells from the gonads of wild-type and SF-1 KO mice, allowing us to compare gene expression profiles of the same cell populations in the presence or absence of SF-I. We will use transient transfection experiments with the 5-flanking regions of these candidate genes to determine if they are direct or indirect targets of SF-1. To develop a mechanism to inactivate the expression of genes of interest from early stages of gonadogenesis, we will use BAC transgenesis to target expression of Cre recombinase to the sites where SF-1 is expressed. Finally, we will attempt to define the sequences that regulate SF-1 expression in the gonads, focusing on a conserved sequence in the 6th intron that is associated with a DNase hypersensitive site. These studies will provide novel insights into how SF-1 exerts its multiple actions in gonadal development and function.